title: Simultaneous determination of D-aspartic acid and D-glutamic acid in rat tissues and physiological fluids using a multi-loop two-dimensional HPLC procedure.
abstract: For a metabolomics study focusing on the analysis of aspartic and glutamic acid enantiomers, a fully automated two-dimensional HPLC system employing a microbore-ODS column and a narrowbore-enantioselective column was developed. By using this system, a detailed distribution of D-Asp and D-Glu besides L-Asp and L-Glu in mammals was elucidated. For the total analysis concept, the amino acids were first pre-column derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) to be sensitively and fluorometrically detected. For the non-stereoselective separation of the analytes in the first dimension a monolithic ODS column (750 mm × 0.53 mm i.d.) was adopted, and a self-packed narrowbore-Pirkle type enantioselective column (Sumichiral OA-2500S, 250 mm × 1.5 mm i.d.) was selected for the second dimension. In the rat plasma, RSD values for intra-day and inter-day precision were less than 6.8%, and the accuracy ranged between 96.1% and 105.8%. The values of LOQ of D-Asp and D-Glu were 5 fmol/injection (0.625 nmol/g tissue). The present method was successfully applied to the simultaneous determination of free aspartic acid and glutamic acid enantiomers in 7 brain areas, 11 peripheral tissues, plasma and urine of Wistar rats. Biologically significant D-Asp values were found in various tissue samples whereas for D-Glu the values were very low possibly indicating less significance.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Fukuoka 812-8582, Japan.
authors.forename: Hai
authors.initials: H
authors.lastname: Han
authors.forename: Yurika
authors.initials: Y
authors.lastname: Miyoshi
authors.forename: Kyoko
authors.initials: K
authors.lastname: Ueno
authors.forename: Chieko
authors.initials: C
authors.lastname: Okamura
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Tojo
authors.forename: Masashi
authors.initials: M
authors.lastname: Mita
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Lindner
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Zaitsu
authors.forename: Kenji
authors.initials: K
authors.lastname: Hamase
mesh_terms: D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives / Q000737:chemistry; D000818:Animals; D001923:Brain Chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D026603:D-Aspartic Acid / Q000032:analysis* / Q000097:blood / Q000737:chemistry / Q000652:urine; D018698:Glutamic Acid / Q000032:analysis* / Q000097:blood / Q000737:chemistry / Q000652:urine; D016014:Linear Models; D008297:Male; D009928:Organ Specificity; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013237:Stereoisomerism
chemical_list: C032930:7-fluoro-4-nitrobenzo-2-oxa-1,3-diazole; D018698:Glutamic Acid; D026603:D-Aspartic Acid; D009327:4-Chloro-7-nitrobenzofurazan